1. Field of the Invention
The present invention provides methods and compositions for identifying human subjects with an increased risk of having essential hypertension. In particular, this invention relates to the identification and characterization of polymorphisms in the human tissue kallikrein gene promoter correlated with increased or decreased risk of developing essential hypertension and the responsiveness of a subject to various treatments for hypertension.
2. Background Art
The tissue kallikrein-kinin system has long been implicated in blood pressure regulation by genetic and physiological studies. Urinary excretion of tissue kallikrein is significantly lower in patients with essential hypertension (Elliot et al., 1934; Carretero et al., 1971; Margolius et al., 1971; Margolius et al., 1974) and in genetically hypertensive rats (Carretero et al., 1976; Carretero et al., 1978; Ader et al., 1986; Ader et al., 1987). Total urinary tissue kallikrein excretion was also found to be lower among individuals with parental hypertension than it was in individuals with two normotensive parents (Williams et al, 1987). Transgenic mice over-expressing the human tissue kallikrein gene have been shown to exhibit a hypotensive phenotype (Wang et al., 1994; Song et al., 1995). Further, a study involving 57 Utah pedigrees indicates that a dominant allele expressed as high total urinary kallikrein excretion may be associated with decreased risk of essential hypertension (Berry et al., 1989).
Identification of DNA polymorphisms provides an efficient way to study the association of genes and diseases by linkage analysis. The human tissue kallikrein gene is located on the long arm of chromosome 19 q13.3-13.4 (Evans et al., 1988). Although a TaqI restriction fragment length polymorphism (RFLP) was found at this locus in a Norwegian population (Berge and Berg, 1991), this genetic marker has only two polymorphisms with unfavored allelic frequencies, making it unsuitable for genetic studies. No significant linkage was found between blood pressure and the kallikrein gene using this marker (Berge and Berg, 1993). A polymorphic microsatellite marker close to the tissue kallikrein gene locus has been identified which is useful for linkage analysis (Richards et al., 1991). However, no previous studies have demonstrated a correlation between specific polymorphisms in the tissue kallikrein gene and blood pressure.
A major challenge in the hypertension field is the development and implementation of rational therapeutic approaches and new preventive measures. High blood pressure is associated with the risk of coronary heart disease and stroke not only among xe2x80x9chypertensivexe2x80x9d patients, but also among xe2x80x9cnormotensivexe2x80x9d individuals (MacMahon et al., 1990). A reduction of 5 mmHg in diastolic blood pressure could result in a 21% reduced risk for coronary heart disease and 34% for stroke (MacMahon et al., 1990). Because a universal salt restriction may be inappropriate at the population level due to the heterogeneity of blood pressure responses (Pecker and Laragh, 1991), the need exists for a marker to predict blood pressure responsive variations with regard to particular clinical treatments. A link between kallikrein promoter polymorphism, urinary kallikrein level, salt sensitivity and blood pressure response would have great utility in categorizing individuals for particular clinical treatments, such as a preferential low salt recommendation for salt-sensitive patients, and a directional selection of anti-hypertensive drugs [e.g., diuretics and angiotension-converting enzyme (ACE) inhibitors] according to a patient""s genotype.
The present invention overcomes previous shortcomings in the art by providing methods and compositions for identifying individuals having an increased or decreased risk of developing essential hypertension and identifying the responsiveness of individuals to certain treatment modalities, based on analysis and characterization of a highly polymorphic locus in the promoter region of the human tissue kallikrein gene.
The present invention provides a method for identifying a human subject as having an increased risk of developing essential hypertension, comprising determining the presence in the subject of an allele in the promoter region of the subject""s tissue kallikrein gene correlated with an increased risk of developing essential hypertension, the presence of the allele identifying the subject as having an increased risk of developing essential hypertension.
Also provided is a method for identifying a human subject as having an increased risk of developing essential hypertension, comprising correlating the presence of a nucleic acid sequence at nucleotides xe2x88x92166 through xe2x88x9298 in a human tissue kallikrein gene with an increased risk of developing essential hypertension; and determining the nucleic acid sequence of nucleotides xe2x88x92166 through xe2x88x9298 of the subject""s tissue kallikrein gene, whereby a subject having a nucleic acid sequence at nucleotides xe2x88x92166 through xe2x88x9298 in the tissue kallikrein gene correlated with an increased risk of developing essential hypertension is identified as having an increased risk of developing essential hypertension.
Furthermore, a method of identifying an allele in the promoter region of the human tissue kallikrein gene correlated with an increased risk of developing essential hypertension is also provided, comprising determining the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298 in the human tissue kallikrein gene from a subject; and correlating the presence of the nucleic acid sequence of the subject with the diagnosis of essential hypertension in the subject, whereby the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298 in the promoter region of the human tissue kallikrein gene identifies an allele correlated with an increased risk of developing essential hypertension.
Additionally provided is a method of identifying an allele in the promoter region of a human tissue kallikrein gene correlated with an increased risk of developing essential hypertension, comprising determining the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298 in the promoter region of the human tissue kallikrein gene; cloning the nucleic acid sequence encoding the promoter region of the human tissue kallikrein gene into an expression vector having a reporter gene; expressing the reporter gene to produce a gene product under the direction of the promoter region of the human tissue kallikrein gene; and determining the amount of reporter gene product produced, an amount of reporter gene product less than the amount of reporter gene product produced under the direction of a tissue kallikrein gene promoter containing the nucleotide sequence of SEQ ID NO:14 (allele A) or the nucleotide sequence of SEQ ID NO:15 (allele B) identifying a nucleic acid sequence of the nucleotides at position xe2x88x92166 through xe2x88x9298 of the human tissue kallikrein gene as an allele correlated with an increased risk of developing essential hypertension.
Further provided is a method for identifying a human subject as having an increased risk of developing essential hypertension, comprising cloning the nucleic acid sequence encoding the promoter region of the human tissue kallikrein gene of the subject into an expression vector having a reporter gene; expressing the reporter gene to produce a gene product under the direction of the promoter region of the human tissue kallikrein gene; and determining the amount of reporter gene product produced, an amount of reporter gene product less than the amount of reporter gene product produced under the direction of a tissue kallikrein gene promoter containing the nucleotide sequence of SEQ ID NO:14 (allele A) or the nucleotide sequence of SEQ ID NO:15 (allele B) identifying a subject as having an increased risk of developing essential hypertension.
In addition, a method for identifying an allele in the promoter region of the human tissue kallikrein gene is provided, comprising determining the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298.
The present invention further provides an isolated nucleic acid consisting of the promoter region of the human tissue kallikrein gene between positions xe2x88x921 to xe2x88x92940, the complementary sequence of the isolated nucleic acid, vectors containing the nucleic acid and cells expressing the vector.
The present invention also provides a method of identifying a subject at reduced risk of having essential hypertension, comprising determining the presence in the subject of an allele in the promoter region of the subject""s tissue kallikrein gene correlated with a reduced risk of developing essential hypertension, the presence of the allele identifying the subject as having a reduced risk of developing essential hypertension.
Additionally provided in the present invention is a method for identifying a human subject as having a reduced risk of developing essential hypertension, comprising correlating the presence of a nucleic acid sequence at nucleotides xe2x88x92166 through xe2x88x9298 in a human tissue kallikrein gene with a reduced risk of developing essential hypertension; and determining the nucleic acid sequence of nucleotides xe2x88x92166 through xe2x88x9298 of the subject""s tissue kallikrein gene, whereby a subject having a nucleic acid sequence at nucleotides xe2x88x92166 through xe2x88x9298 in the tissue kallikrein gene correlated with a reduced risk of developing essential hypertension is identified as having a reduced risk of developing essential hypertension.
A method is also provided for identifying an allele in the promoter region of the human tissue kallikrein gene correlated with a reduced risk of developing essential hypertension comprising determining the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298 in the human tissue kallikrein gene from a subject; and correlating the presence of the nucleic acid sequence of the subject with the absence of essential hypertension in the subject, whereby the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298 in the promoter region of the human tissue kallikrein gene identifies an allele correlated with a reduced risk of developing essential hypertension.
The present invention further provides a method of identifying an allele in the promoter region of a human tissue kallikrein gene correlated with a reduced risk of developing essential hypertension, comprising determining the nucleic acid sequence of the nucleotides at positions xe2x88x92166 through xe2x88x9298 in the promoter region of the human tissue kallikrein gene; cloning the nucleic acid sequence encoding the promoter region of the human tissue kallikrein gene into an expression vector having a reporter gene; expressing the reporter gene to produce a gene product under the direction of the promoter region of the human tissue kallikrein gene; and determining the amount of reporter gene product produced, an amount of reporter gene product greater than the amount of reporter gene product produced under the direction of a tissue kallikrein gene promoter containing the nucleotide sequence of SEQ ID NO:14 (allele A) or the nucleotide sequence of SEQ ID NO:15 (allele B) identifying a nucleic acid sequence of the nucleotides at position xe2x88x92166 through xe2x88x9298 of the human tissue kallikrein gene as an allele correlated with a reduced risk of developing essential hypertension.
Also provided is a method for identifying a human subject as having a reduced risk of developing essential hypertension, comprising cloning the nucleic acid sequence encoding the promoter region of the human tissue kallikrein gene of the subject into an expression vector having a reporter gene; expressing the reporter gene to produce a gene product under the direction of the promoter region of the human tissue kallikrein gene; and determining the amount of reporter gene product produced, an amount of reporter gene product greater than the amount of reporter gene product produced under the direction of a tissue kallikrein gene promoter containing the nucleotide sequence of SEQ ID NO:14 (allele A) or the nucleotide sequence of SEQ ID NO:15 (allele B) identifying a subject as having a reduced risk of developing essential hypertension.
In addition to the embodiments described above, the present invention provides a method of identifying a human subject as having an increased likelihood of regulating blood pressure with dietary sodium intake, comprising determining the presence in the subject of a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with dietary sodium intake, whereby the presence of the genotype identifies the subject as having an increased likelihood of regulating blood pressure by dietary sodium intake.
Additionally provided in this invention is a method of identifying a human subject as having an increased likelihood of regulating blood pressure with dietary sodium intake, comprising a) correlating the presence of a tissue kallikrein promoter allele genotype with an increased likelihood of regulating blood pressure with dietary sodium intake; and b) determining the tissue kallikrein promoter allele genotype of the subject, whereby a subject having a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with dietary sodium intake is identified as having an increased likelihood of regulating blood pressure with dietary sodium intake.
Furthermore, a method is provided for identifying a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with dietary sodium intake, comprising a) determining the tissue kallikrein promoter allele genotype of a subject; and b) correlating the presence of the genotype of step (a) with the ability of the subject to regulate blood pressure by dietary sodium intake, thereby identifying a genotype correlated with an increased likelihood of regulating blood pressure by dietary sodium intake.
Additionally, the present invention provides a method for identifying a human subject as having a reduced likelihood of regulating blood pressure with dietary sodium intake, comprising determining the presence in the subject of a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with dietary sodium intake, whereby the presence of the genotype identifies the subject as having a reduced likelihood of regulating blood pressure with dietary sodium intake.
Additionally provided in this invention is a method of identifying a human subject as having a reduced likelihood of regulating blood pressure with dietary sodium intake, comprising a) correlating a tissue kallikrein promoter allele genotype with a reduced likelihood of regulating blood pressure with dietary sodium intake; and b) determining the tissue kallikrein promoter allele genotype of the subject, whereby a subject having a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with dietary sodium intake is identified as having a reduced likelihood of regulating blood pressure with dietary sodium intake.
Also, a method is provided of identifying a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with dietary sodium intake, comprising a) determining the tissue kallikrein promoter allele genotype of a subject; and b) correlating the presence of the genotype of step (a) with the inability of the subject to regulate blood pressure by dietary sodium intake, thereby identifying the genotype correlated with a reduced likelihood of regulating blood pressure by dietary sodium intake.
In a further embodiment, the present invention provides a method of identifying a human subject as having an increased likelihood of regulating blood pressure with a diuretic, comprising determining the presence in the subject of a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with a diuretic, whereby the presence of the genotype identifies the subject as having an increased likelihood of regulating blood pressure with a diuretic.
Additionally provided is a method of identifying a human subject as having an increased likelihood of regulating blood pressure with a diuretic, comprising a) correlating the presence of a tissue kallikrein promoter allele genotype with an increased likelihood of regulating blood pressure with a diuretic; and b) determining the tissue kallikrein promoter allele genotype of the subject, whereby a subject having a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with a diuretic is identified as having an increased likelihood of regulating blood pressure with a diuretic.
Also provided herein is a method of identifying a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with a diuretic, comprising a) determining the tissue kallikrein promoter allele genotype of a subject; and b) correlating the presence of the genotype of step (a) with the ability of the subject to regulate blood pressure with a diuretic, thereby identifying a genotype correlated with an increased likelihood of regulating blood pressure with a diuretic.
A method of identifying a human subject as having a reduced likelihood of regulating blood pressure with a diuretic is also provided, comprising determining the presence in the subject of a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with a diuretic, whereby the presence of the genotype identifies the subject as having a reduced likelihood of regulating blood pressure with a diuretic.
Furthermore, the present invention provides a method of identifying a human subject as having a reduced likelihood of regulating blood pressure with a diuretic, comprising a) correlating a tissue kallikrein promoter allele genotype with a reduced likelihood of regulating blood pressure with a diuretic; and b) determining the tissue kallikrein promoter allele genotype of the subject, whereby a subject having a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with a diuretic is identified as having a reduced likelihood of regulating blood pressure with a diuretic.
Also provided is a method of identifying a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with a diuretic, comprising a) determining the tissue kallikrein promoter allele genotype of a subject; and b) correlating the presence of the genotype of step (a) with the inability of the subject to regulate blood pressure with a diuretic, thereby identifying the genotype as correlated with a reduced likelihood of regulating blood pressure with a diuretic.
The present invention additionally provides a method of identifying a human subject as having an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, comprising determining the presence in the subject of a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, whereby the presence of the genotype identifies the subject as having an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor.
Furthermore, the present invention provides a method of identifying a human subject as having an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, comprising a) correlating the presence of a tissue kallikrein promoter allele genotype with an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor; and b) determining the tissue kallikrein promoter allele genotype of the subject, whereby a subject having a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor is identified as having an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor.
Also provided herein is a method of identifying a tissue kallikrein promoter allele genotype correlated with an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, comprising a) determining the tissue kallikrein promoter allele genotype of a subject; and b) correlating the presence of the genotype of step (a) with the ability of the subject to regulate blood pressure with an angiotensin-converting enzyme inhibitor, thereby identifying a genotype correlated with an increased likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor.
A method of identifying a human subject as having a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor is additionally provided, comprising determining the presence in the subject of a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, whereby the presence of the genotype identifies the subject as having a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor.
In addition to the above embodiments, the present invention also provides a method of identifying a human subject as having a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, comprising a) correlating a tissue kallikrein promoter allele genotype with a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor; and b) determining the tissue kallikrein promoter allele genotype of the subject, whereby a subject having a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor is identified as having a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor.
Finally, the present invention provides a method of identifying a tissue kallikrein promoter allele genotype correlated with a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor, comprising a) determining the tissue kallikrein promoter allele genotype of a subject; and b) correlating the presence of the genotype of step (a) with the inability of the subject to regulate blood pressure with an angiotensin-converting enzyme inhibitor, thereby identifying the genotype as correlated with a reduced likelihood of regulating blood pressure with an angiotensin-converting enzyme inhibitor.
Various other objectives and advantages of the present invention will become apparent from the following description.